The present invention relates to a diagnostic method to monitor coreceptor use in treatment of human immunodeficiency virus (HIV, or xe2x80x9can AIDS virusxe2x80x9d) infection. This method can be used to assist in selecting antiretroviral therapy and to improve predictions of disease prognosis. The present invention further relates to a diagnostic method to monitor the suppression of CXCR4-specific strains in HIV infected individuals undergoing antiretroviral therapy. Other aspects of the invention are described in or are obvious from the following disclosure (and within the ambit of the invention).
HIV uses a receptor-mediated pathway in the infection of host cells. HIV-1 requires contact with two cell-surface receptors to gain entry into cells and initiate infection; CD4 is the primary receptor. CXCR4 and CCR5, members of the chemokine receptor family of proteins, serve as secondary coreceptors for HIV-1 isolates that are tropic for T-cell lines or macrophages, respectively. Deng et al. (1996) Nature 381:661-6; Doranz et al. (1996) Cell 86:1149-59; and Berger et al. (1998) Nature 391:240. CXCR4 or CCR5, in conjunction with CD4, form a functional cellular receptor for entry of certain strains of HIV into cells. Recent reports indicated that the viral envelope glycoprotein gp120 interacts directly with chemokine receptors generally at a step following CD4 binding. Lapham et al. (1996) Science 274:602-605; Moore (1997) Science 276:51; Wu et al. (1996) Nature 384:179-183; and Hesselgesser et al. (1997) Current Biology 7:112-121. Envelope variants will selectively interact with either CXCR4 or CCR5.
HIV-1 strains transmitted in vivo generally use CCR5 (CCR5 viruses). Fenyo et al. (1998) Nature 391:240; Samson et al. (1996) Nature 382:722-5; Shankarappa et al. (1999) J. Virol. 73:10489-502; and Scarlatti et al. (1997) Nature Med. 3:1259-65. These viruses typically infect macrophages and primary CD4+ lymphocytes, and do not form syncytia in vitro. Bjxc3x6rndal et al. (1997) J. Virol. 71:7478-87. These viruses are said to be macrophage tropic (M-tropic). After primary HIV-1 infection, viral populations are usually characterized by molecular heterogeneity. Shankarappa et al. (1999); and Glushakova et al. (1999) J. Clin. Invest. 104:R7-R11.
Years after chronic infection is established, strains using CXCR4 emerge in xcx9c50% of infected individuals. Berger et al. (1998); Scarlatti et al. (1997); Koot et al. (1993); and Connor et al. (1997) J. Exp. Med. 185:621-8. CXCR4 strains not only infect primary T-lymphocytes but also replicate in T-cell lines and induce syncytia. Bjxc3x6rndal et al. (1997). These viruses are said to be T-cell tropic (T-tropic). This difference in cell tropism correlates with disease progression. During HIV infection, strains isolated from individuals early in the course of their infection are usually M-tropic, while viruses isolated from approximately 50% of individuals with advanced immunodeficiency also include viruses that are T-tropic.
The finding that change from M- to T-tropic viruses over time in infected individuals correlates with disease progression suggested that the ability of the viral envelope to interact with CXCR4 represents an important feature in the pathogenesis of immunodeficiency and the development of full blown Acquired Immunodeficiency Syndrome (AIDS).
CXCR4 strains have now been shown to have a striking influence on HIV-1 disease progression. Cytopathicity toward the general CD4+ T cell population in lymphoid tissue is associated with the use of CXCR4. Glushakova et al. (1999). The emergence of CXCR4 virus is predictive of rapid depletion of CD4+ cells and acceleration of HIV-1 disease progression. Berger et al. (1998); Scarlatti et al. (1997); Koot et al. (1993); and Connor et al. (1997). A recent analysis of HIV-1 coreceptor use in infected individuals suggested that the rapid CD4+ cell decline is related to the ability of CXCR4 viruses to infect an expanded spectrum of crucial target cells as compared to CCR5 strains. Blaak et al. (2000) Proc. Natl. Acad. Sci. USA 97:1269-74. In vitro results suggest that selective blockade of CXCR4 receptors may prevent the switch from the less pathogenic CCR5 strains to the more pathogenic CXCR4 strains. Este et al. (1999) J. Virol. 73:5577-85. Coreceptor use plays a critical role in viral tropism, pathogenesis, and disease progression. Thus, a diagnosic method for use in detecting CXCR4 isolates and/or monitoring shifts in coreceptor use would be beneficial for predicting disease progression over time.
Treatment of infected individuals with highly active antiretroviral therapy (HAART) has led to a dramatic decline in both HIV-1-related illness and death. Palella et al. (1998) N. Engl. J. Med. 338:853-60. Early clinical trials demonstrated a reduction of plasma HIV-1 RNA loads to undetectable levels in the majority of treated individuals. Hammer et al. (1997) N. Engl. J. Med. 337:725-33; and Autran et al. (1997) Science 277:112-6. Subsequent studies, however, showed more limited success in achieving and maintaining viral suppression. Deeks et al. (2000) J. Inf. Dis. 181:946-53; and Mezzaroma et al. (1999) Clin. Inf. Dis. 29:1423-30. Yet many patients experienced immunologic and clinical responses to HAART without sustained suppression of plasma viremia. Deeks et al. (2000); and Mezzaroma et al. (1999).
The emergence of viral variants in connection with the failure of HAART may be associated with modified expression of the host determinants of viral tropism, including CCR5 and CXCR4. In comparison to pretherapy determinations, expression of CXCR4 was significantly increased, and CCR5 decreased, following three months of an anti-viral regimen. Giovannetti et al. (1999) Clin. Exp. Immunol. 118:87-94. Changes in coreceptor expression occurred in association with a decrease in viral load and T cell activation, and an increase in naive and memory T cells signifying peripheral redistribution of T cell compartments. In a separate study, HAART was reported to reduce the expression of CXCR4 and CCR5 in lymphoid tissue. Andersson et al. (1998) AIDS 12:F123-9. These studies did not address coreceptor usage in patients undergoing HAART. The effects of HAART on coreceptor usage by viral populations were heretofore unknown.
Unexpectedly, it has now been shown that in patients undergoing HAART, the predominant populations of virus can be shifted back to CCR5-mediated entry after the CXCR4-specific strains have emerged. Thus, a diagnostic method for use in monitoring shifts in coreceptor use would be beneficial for measuring the therapeutic efficacy of various HIV treatment regimes, such as HAART.
The correlation between CXCR4-specific strains and rapid disease progression indicates that a diagnostic method would be useful to monitor the presence of CXCR4-specific strains and shifts in coreceptor use associated with HIV disease progression. Application of the diagnostic method allows more accurate predictions of disease prognosis over time.
The effect of HAART on coreceptor use by populations of virus has not heretofore been quantitatively studied. Herein, it is shown that in patients undergoing combination antiretroviral therapy, including HAART, the predominant populations of virus can be shifted back to CCR5-mediated entry once the CXCR4-specific strains have emerged.
Therefore, a diagnostic method is also useful to monitor the presence of CXCR4-specific strains and shifts in coreceptor use in patients undergoing antiretroviral therapy. Application of the diagnostic method allows the effectiveness of antiretroviral therapy to be more closely monitored.
The present invention relates to a diagnostic method to determine whether CXCR4 or CCR5 isolates are present in a patient comprising assaying for coreceptor use.
The present invention further relates to a diagnostic method to determine whether CXCR4 or CCR5 strains are present in a patient comprising transforming cells with an HIV envelope gene variant cloned from an infected patient, selectively fusing the cells with an indicator cell line expressing an HIV envelope-compatible coreceptor, and assaying for fusion.
The present invention further relates to a diagnostic method to determine whether CXCR4 or CCR5 strains are present in a patient comprising obtaining patient-derived virus and assaying the isolates for coreceptor use.
The present invention further relates to a diagnostic method to monitor shifts in coreceptor use associated with changes in HIV disease progression.
The present invention further relates to a diagnostic method to monitor shifts in coreceptor use associated with changes in HIV disease progression comprising transforming cells with an HIV envelope gene variant cloned from an infected patient, selectively fusing the cells with an indicator cell line expressing an HIV envelope-compatible coreceptor, and assaying for fusion.
The present invention further relates to a diagnostic method to monitor shifts in coreceptor use associated with changes in HIV disease progression comprising transforming cells containing an HIV Tat-activatable reporter gene construct with an HIV envelope gene variant cloned from an infected patient, selectively fusing the cells with an indicator cell line containing a constitutively active tat gene and an HIV envelope-compatible coreceptor, and assaying for fusion by detection of reporter gene expression.
The present invention further relates to a diagnostic method to monitor shifts in coreceptor use associated with changes in HIV disease progression comprising determining CXCR4 coreceptor use, CCR5 coreceptor use, and a ratio of HIV using the CXCR4 coreceptor compared to HIV using the CCR5 coreceptor.
The present invention further relates to a diagnostic method to monitor shifts in coreceptor use associated with changes in HIV disease progression by obtaining patient-derived virus and deriving biological clones therefrom, assaying the clones for coreceptor use, and applying a method of quantitating the proportion of virus that uses each coreceptor.
The present invention further relates to a diagnostic method to determine whether CXCR4 or CCR5 strains are present in patients infected with HIV undergoing antiretroviral therapy.
The present invention further relates to a diagnostic method to determine whether CXCR4 or CCR5 strains are present in patients infected with HIV undergoing antiretroviral therapy comprising transforming cells with an HIV envelope gene variant cloned from an infected patient, selectively fusing the cells with an indicator cell line expressing an HIV envelope-compatible coreceptor, and assaying for fusion before and/or after initiating antiretroviral therapy.
The present invention further relates to a diagnostic method to determine whether CXCR4 or CCR5 strains are present in patients infected with HIV undergoing antiretroviral therapy comprising transforming cells containing an HIV Tat-activatable reporter gene construct with an HIV envelope gene variant cloned from an infected patient, selectively fusing the cells with an indicator cell line containing a constitutively active tat gene and an HIV envelope-compatible coreceptor, and assaying for fusion by detection of reporter gene expression before and/or after initiating antiretroviral therapy.
The present invention further relates to a diagnostic method to determine whether CXCR4 or CCR5 strains are present in patients infected with HIV undergoing antiretroviral therapy comprising obtaining patient-derived virus and assaying the virus for coreceptor use before and/or after initiating antiretroviral therapy.
The present invention further relates to a diagnostic method to monitor the suppression of CXCR4-specific strains in patients infected with HIV undergoing antiretroviral therapy.
The present invention further relates to a diagnostic method to monitor the suppression of CXCR4-specific strains in patients infected with HIV undergoing antiretroviral therapy comprising transforming cells with an HIV envelope gene variant cloned from an infected patient, selectively fusing the cells with an indicator cell line expressing an HIV envelope-compatible coreceptor, and assaying for fusion before and/or after initiating antiretroviral therapy.
The present invention further relates to a diagnostic method to monitor the suppression of CXCR4-specific strains in patients infected with HIV undergoing antiretroviral therapy comprising transforming cells containing an HIV Tat-activatable reporter gene construct with an HIV envelope gene variant cloned from an infected patient, selectively fusing the cells with an indicator cell line containing a constitutively active tat gene and an HIV envelope-compatible coreceptor, and assaying for fusion by detection of reporter gene expression before and/or after initiating antiretroviral therapy.
The present invention further relates to a diagnostic method to monitor the suppression of CXCR4-specific strains in patients infected with HIV undergoing antiretroviral therapy comprising obtaining patient-derived virus and assaying the virus for coreceptor use before and/or after initiating antiretroviral therapy.
The present invention further relates to a diagnostic method to monitor the suppression of CXCR4-specific strains in patients infected with HIV undergoing antiretroviral therapy comprising determining CXCR4 coreceptor use, CCR5 coreceptor use, and a ratio of HIV using the CXCR4 coreceptor compared to HIV using the CCR5 coreceptor before and/or after initiating antiretroviral therapy.
The present invention further relates to a diagnostic method to monitor the suppression of CXCR4-specific strains in patients infected with HIV undergoing antiretroviral therapy comprising obtaining primary viral isolates and deriving biological clones therefrom, assaying the clones for coreceptor use, and applying a method for quantitating the proportion of virus in a primary isolate that uses each coreceptor before and/or after initiating antiretroviral therapy.
The present invention further relates to a diagnostic method to monitor the suppression of CXCR4-specific strains in patients infected with HIV undergoing antiretroviral therapy comprising determining the sequence of HIV envelope gene before and/or after initiating antiretroviral therapy.
These and other objects and embodiments are described in or are obvious from and within the scope of the invention, from the following Detailed Description.